Process for preparation of isosulfan blue

ABSTRACT

A process for the preparation of isosulfan blue (Active Pharmaceutical Ingredient) is provided. A process is also provided for preparation of the intermediate, 2-chlorobenzaldehyde-5-sulfonic acid, sodium salt of formula (2), used in the preparation thereof and a procedure for the isolation of benzaldehyde-2,5-disulfonic acid, di-sodium salt of the formula (3). Also provided is a process for the preparation of an isoleuco acid of formula (4), which upon mild oxidation gives rise to isosulfan blue of pharmaceutical grade which can be used for preparation of pharmaceutical formulations. The isolation and purification procedures provided in the process provide substantially pure isosulfan blue with HPLC purity 99.5% or greater.

CROSS-REFERENCE TO RELATED APPLICATIONS

This is a continuation application, and claims the benefit, of U.S.patent application Ser. No. 13/310,019, filed Dec. 2, 2011, which is acontinuation of U.S. patent application Ser. No. 12/643,056, filed Dec.21, 2009, now abandoned, which is a continuation of U.S. Ser. No.12/180,057 filed Jul. 25, 2008, now U.S. Pat. No. 7,662,992, which is acontinuation of U.S. Ser. No. 11/747,291 filed May 11, 2007, nowabandoned, the entireties of which are incorporated herein by reference.

FIELD OF THE INVENTION

The present invention relates to a process for the production ofisosulfan blue, and in particular, to a process for the production ofisosulfan blue in a substantially pure form.

BACKGROUND OF THE INVENTION

Isosulfan blue, having a chemical name, N-[4-[[4-(diethyl amino)phenyl](2,5-disulfophenyl)methylene]-2,5-cyclohexadien-1-ylidene]-N-ethylethanaminium, sodium saltand the formula

is a triarylmethane dye used as a contrast agent for the delineation oflymphatic vessels and is particularly useful as a cancer diagnosticagent. Also known chemically as sulfan blue or patent blue, isosulfanblue is an active pharmaceutical ingredient used in the Lymphazurin™blue dye pharmaceutical dosage form, available as 1% (10 mg/ml) 5 mlsolution in phosphate buffer for injection. It is commonly used in aprocedure called “mapping of the sentinel lymph nodes”. It is an adjunctto lymphography for visualization of the lymphatic system draining theregion of injection. It has been used with increasing frequency inlocalizing sentinel lymph nodes in breast cancer patients. Isosulfanblue-guided surgical removal of cancerous tissue has been on the rise asit is cost effective and safer to use than technetium 99Mradioisotope-labeled sulfur colloid. Isosulfan blue is a structuralisomer of sulphan blue; both belong to the family of triarylmethanedyestuffs. Generally, preparation of triarylmethane dyes involvescondensation of suitably substituted aryl aldehydes with 2 equivalentsof alkyl-aryl amines giving rise to leuco-bases or leuco-acids followedby oxidation. Although the literature is replete with methods ofpreparing triarylmethane dyes, most of the methods involve strong acidsfor condensation resulting in leuco-bases or leuco-acids, hazardousoxidizing agents (lead oxide, chloranil, iron phthalocyanine/oxone) forconverting to triarylmethane dyes, and crude methods (precipitation withsodium sulfate) of purification. See for example U.S. Pat. Nos.4,330,476, 4,710,322, 1,531,507, 5,659,053, 1,805,925, 2,422,445,1,878,530 and 2,726,252. Prior art methods of isolation of the crudeleuco-acids or leuco-bases involve tedious neutralization/basificationwith strong bases and typically using the reaction mixtures in theoxidation step, giving rise to crude triarylmethane dyes. Thetriarylmethane dyestuffs thus prepared are used mainly for dyeingfabric, coloring paper, and printing inks The literature citesutilization of the same aforementioned synthetic and isolation methodsfor the preparation of diagnostically important dyes, such as isosulfanblue, sulphan blue and patent blue V. See, Rodd's Chemistry of CarbonCompounds by S. Coffey, 1974 2^(nd) Edition, Volume III Part F, 110-133.

Therefore there is a need in the art for an improved method in theprocess chemistry of isosulfan blue to be prepared in the purest formwhich is suitable for large scale cGMP production for its pharmaceuticalformulation manufacturing.

SUMMARY OF THE INVENTION

It is therefore an object of the present invention is to provide asimple, safe, cost-effective, time saving and reliable process for thepreparation of isosulfan blue in bulk scale and in substantially pureform. “Substantially pure” is defined herein as 99.0% or greater.

Another object of the invention is to provide a simple, cost-effectiveand reliable process for preparation of the intermediate,2-chlorobenzaldehyde-5-sulfonic acid, sodium salt of formula (2),required in the preparation of isosulfan blue. This embodiment providesa process step that does not require tedious neutralization with verylarge quantities of sodium carbonate and effervescence, as is the casein prior art processes.

Another object of the invention is to provide a simplified procedure forthe isolation of benzaldehyde-2,5-disulfonic acid, di-sodium salt of theformula (3) that does not include acidifying the reaction mixture withconcentrated sulfuric acid and boiling until excess sulfurous acid isexpelled, as is taught in the prior art.

Yet another object of the invention is to provide a procedure forobtaining the benzaldehyde-2,5-disulfonic acid, sodium salt of formula(3) free of inorganic salts, which essentially simplifies the isolationprocedures to be implemented during isolation of isoleuco acid.

Yet another, object of the invention is to provide a process for thepreparation of an isoleuco acid of formula (4), through the ureaderivative as an in-situ intermediate. The isoleuco acid of formula (4)on further oxidation gives rise to the target compound, isosulfan blue(5). Still another object of the invention is to use very mild oxidationagent to avoid any over oxidized products and also to improve thestability of the isosulfan blue under reaction conditions.

According to this invention, there is provided a simple procedure forthe isolation of benzaldehyde-2,5-disulfonic acid, isoleuco acid andisosulfan blue at acid stage and also at sodium salt formation stage byincorporating crystallization techniques, thereby avoiding distillationand other techniques using high temperatures which jeopardize thecompound stability during the manufacturing process.

These and other aspects of the invention will be apparent to thoseskilled in the art.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

In the following description, for purposes of explanation, specificnumbers, materials and configurations are set forth in order to providea thorough understanding of the invention. It will be apparent, however,to one having ordinary skill in the art that the invention may bepracticed without these specific details. In some instances, well-knownfeatures may be omitted or simplified so as not to obscure the presentinvention. Furthermore, reference in the specification to phrases suchas “one embodiment” or “an embodiment” means that a particular feature,structure or characteristic described in connection with the embodimentis included in at least one embodiment of the invention. The appearancesof phrases such as “in one embodiment” in various places in thespecification are not necessarily all referring to the same embodiment.In accordance with one embodiment the present invention relates to aprocess for the preparation of isosulfan blue.

Experimental Procedures

In accordance with one embodiment of the present invention a first stepinvolves sulfonation of the commercially available starting material ofthe formula (1) to 2-chlorobenzaldehyde-5-sulfonic acid sodium salt ofthe formula (2).

In one example, the sulfonation process involved reacting one equivalentof the 2-chlorobenzaldehyde of formula (1) with 2.0 equivalents of 20%fuming sulfuric acid at 15° C. to 70° C. for 16 hrs. The reactionmixture was poured into ice-water carefully followed by stirring withsolid sodium chloride resulting in a cream colored precipitate, whichupon filtration, washing with ether and drying afforded2-chlorobenzaldehyde-5-sulfonic acid of the formula (2) in 86% yield.

In accordance with one embodiment of the present invention, a secondstep of the process involves nucleophilic displacement of the chloridein 2-chlorobenzaldehyde-5-sulfonic acid sodium salt of the formula (2)with an alkali metal sulfite/bisulfite such as sodium sulfite/sodiumbisulfite at elevated temperatures under closed conditions.

In one example, this reaction was carried out in a Parr pressure vesselequipped with overhead magnetic stirring.2-Chlorobenzaldehyde-5-sulfonic acid (2), sodium sulfite (2.29equivalents), sodium bisulfite (10% of sodium sulfite), and water (3.45mL/g) were charged into the Parr pressure vessel. The reaction mixturein the vessel was stirred and heated at 170-180° C. for 5-7 hoursgenerating 140-150 psi pressure.

The reaction mixture, after cooling, was poured into methanol whilestirring, so as to make 20% aqueous content of the whole volume. Thisprocess ensured total precipitation of the inorganic salts, which couldbe removed by filtration. The solvent from the filtrate was removedunder reduced pressure to obtain a solid residue, which was trituratedwith methanol and filtered to afford light yellow colored compound,benzaldehyde-2,5-disulfonic acid, di sodium salt of the formula (3) in93.9% yield.

In accordance with one embodiment a purification procedure for removingthe inorganic salts essentially involves dissolving the crude solid inN,N dimethylformamide and stirring the contents for 1-2 hours at ambienttemperature followed by filtration. The filtrate is precipitated bydichloromethane to afford the light yellow colored compound,benzaldehyde-2,5-disulfonic acid disodium salt of formula (3) withchromatographic purity NLT 99.0% and with HPLC assay greater than 90%w/w.

In accordance with one embodiment of the present invention, a third stepof the process involved condensing benzaldehyde-2,5-disulfonic acid,disodium salt of the formula (3) with N, N-diethylaniline to provideisoleuco acid of the formula (4).

In one example, pure isoleuco-acid of the formula (4) withchromatographic purity greater than 98.0% was obtained in the solid formout of the reaction mixture. A mixture of benzaldehyde-2,5-disulfonicacid, disodium salt of the formula (3), N,N-diethylaniline (2.2equivalents), and urea (0.75 equivalents) in glacial acetic acid wasstirred and refluxed for 20-25 hrs. The reaction progressed through theintermediate formation in-situ which is a urea derivative ofbenzaldehyde-2,5-disulfonic acid disodium salt. To the above cooledreaction mixture after 20-25 hrs reflux, methanol was added to form aprecipitate, which was collected by vacuum filtration and washed withdiethyl ether to afford the isoleuco acid of the formula (4) in 56.8%yield.

The purification of isoleuco acid was carried out by dissolving thecrude solid in 5 volumes of water and stirred for 1-2 hours at ambienttemperature and filtering the solid. The above process was repeatedtwice before the final solid was washed with acetone to generateisoleuco acid of the formula (4) with chromatographic purity greaterthan 99.5%.

In accordance with one embodiment of the present invention a fourth stepof the process involves conversion of the isoleuco acid (4) to isosulfanblue of the formula (5) under conditions that employ milder oxidizingagents with no strong acidic reagents and are less hazardous than theprior art.

In an example of the present inventive process, a suspension of isoleucoacid of the formula (4) in methanol was stirred at room temperature for12-14 hrs with silver oxide (2.5 equivalents). The blue colored reactionmixture was filtered through a pad of silica gel and Celite followed byfiltration through an acidic zeolite bed and further through a 0.2micron membrane filtration unit. The filtrate was then precipitated withisopropyl ether at room temperature to obtain crude isosulfan blue acid.

The isosulfan blue acid thus obtained was then purified byrecrystallization from aqueous isopropyl alcohol/acetone to affordisosulfan blue acid of chromatographic purity NLT 99.5% performed byHigh Performance Liquid Chromatography.

The final product of isosulfan blue sodium (formula 5) was obtained whenisosulfan blue acid was adjusted to a pH greater than 6.0 in aqueousacetone medium using sodium bicarbonate solution for pH adjustment. Thereaction mass was filtered to give isosulfan blue sodium of formula (5)having purity greater than 99.5% by HPLC and also free of silver withsilver content estimated by Atomic absorption spectrometer less than 20ppm.

EXAMPLES 2-Chlorobenzaldehyde-5-sulfonic acid, Sodium Salt of theFormula (2)

113.82 g (based on SO₃ molecular weight, 569 mL) of 20% fuming sulfuricacid (FSA) was charged into a 1 L three-neck flask fitted with adropping funnel, overhead stirrer, and thermometer. The reaction masswas cooled to 15 to 20° C. 100 g of 2-chlorobenzaldehyde of the formula(1) was added drop-wise to the stirred and cooled FSA over a period of40 minutes, so that the temperature didn't rise above 20° C. Thereaction mixture was stirred and heated at 70° C. for 16 hours to obtaina dark-brown colored reaction solution. The HPLC results indicated theabsence of the starting material. The dark-brown colored reactionsolution was carefully poured into a beaker containing 1200 g of crushedice and stirred. 500 g of solid sodium chloride was added portion wiseto the stirred colored acidic solution to precipitate a light-yellowcolored solid. The light-yellow colored solid was collected by vacuumfiltration and washed with diethyl ether to afford 150.0 g (86.92%) of2-chlorobenzaldehyde-5-sulfonic acid, sodium salt of the formula (2).

Benzaldehyde-2,5-disulfonic acid, Sodium Salt of the Formula (3)

50 g (0.206 mol) of 2-chlorobenzaldehyde-5-sulfonic acid, sodium salt ofthe formula (2), 59.75 g (0.474 mol, 2.3 eq.) of Na₂SO₃ and 5.97 g (10%of Na₂SO₃) of NaHSO₃ were dissolved in 400 mL of water. The solution wascharged into a 600 mL capacity Parr pressure cylinder equipped withstirring and heating. The reaction mixture was stirred (300-310 RPM) andheated at 180° C. (generates ˜150 psi pressure) for 5-7 hours. HPLCresults indicate the absence of the starting material. After cooling andreleasing the pressure, the reaction mixture was poured into 1600 mL ofstirred methanol and stirred for 15-30 minutes to precipitate theunwanted inorganic salts. The inorganic salts were filtered off using apad of Celite and the filtrate evaporated under reduced pressure toobtain a solid residue. The solid residue obtained was triturated with200 mL methanol, collected by filtration and washed with ether to give60 g (93.9%) of benzaldehyde-2,5-disulfonic acid, sodium salt of formula(3).

Purification of Benzaldehyde-2,5-disulfonic acid, Sodium Salt Formula(3)

60 g of crude benzaldehyde-2,5-disulfonic acid, disodium salt preparedas per the procedure above was dissolved in 500 mL ofN,N-dimethylformamide and stirred for 2 hours at 20-25° C. The mixturewas filtered through a buchner funnel and the filtrate was precipitatedusing 1500 mL of dichloromethane to afford 20 g of the light yellowcolored compound, benzaldehyde-2,5-disulfonic acid disodium salt offormula (3) with chromatographic purity NLT 99.0% w/w.

Isoleuco Acid of the Formula (4)

60 g of benzaldehyde-2,5-disulfonic acid sodium salt of formula (3),8.76 g of urea (0.75 eq), and 1000 mL of glacial acetic acid werecharged into a 3 L 3-neck flask fitted with a mechanical stirrer andreflux condenser. 65.61 mL (2.2 eq) of N,N-diethyl aniline was added tothe stirred mixture and refluxed for 20-25 hrs. When the HPLC resultsindicated the content of starting material was less than 5%, thereaction mass was cooled to room temperature. After cooling to roomtemperature, 600 mL of methanol was added and the separated solidcollected on a sintered funnel by vacuum filtration. The collected solidwas washed with methanol to obtain 55-60 g (56.8%) of crude isoleucoacid of the formula (4).

Purification of Isoleuco Acid of Formula (4)

50 g of crude isoleuco acid along with 250 ml of water was charged intoa 1 L 3-neck round bottom flask fitted with a mechanical stirrer. Thereaction mixture was stirred for 1 hour at 20-25° C. The solid wasfiltered through a buchner funnel.

The above process was repeated twice. The final product thus obtainedwas then washed with 25 ml of acetone and then dried to obtain 40-45 gof the desired isoleuco acid of formula (4).

Isosulfan Blue of the Formula (5)

15 g (0.027 mol) of isoleuco acid of the formula (4) and 225 mL ofMethanol were charged into a 1 L round bottomed flask and the suspensionwas stirred. To the stirred suspension, 15.91 g (0.068 mol, 2.5 eq.) ofsilver oxide was added in one portion at room temperature and stirred atroom temperature for 12-14 hours. The reaction mixture turned blue incolor as the oxidation to the desired product progressed. The HPLCresults indicated the absence of starting material. The blue coloredreaction mixture was filtered through a buchner funnel and the solidsilver oxide collected was taken into the reaction flask and thefiltrate was kept aside. 225 ml of methanol was added to the silveroxide taken in the reaction flask and stirred at 20-25° C. for 30minutes and filtered through the buchner funnel. This silver oxidewashing procedure with methanol was carried out twice more.

The combined filtrates along with the initial filtrate were thenfiltered through a bed of silica gel/celite (2 inch silica gel/1 inch ofcelite) and finally the bed was washed with 50 mL of methanol.

The filtrate was then subjected to a filtration through an acidiczeolite bed of 2 inch height (pH of the zeolite bed was adjusted toacidic pH by using 0.1N hydrochloric acid aqueous solution) followed byfiltration through a 0.2 micron filtration unit.

Isopropyl ether was added three times the volume of the filtrate and theisosulfan blue acid was precipitated as a solid at about 10 gram (68.8%)yield.

In order to prepare the Isosulfan blue sodium salt of the formula (5),10.0 g of the solid obtained above was dissolved in 30 mL deionizedwater. Saturated sodium bicarbonate solution was added drop wise toadjust the pH to 8.0. To this 300 mL of acetone was added and stirred at20-25° C. for 30 minutes. The crystallized product was then filteredthrough a buchner funnel and the solid thus obtained was dried at 40° C.under vacuum to obtain the isosulfan blue sodium salt of formula (5).

While the preferred embodiments have been described and illustrated itwill be understood that changes in details and obvious undisclosedvariations might be made without departing from the spirit and principleof the invention and therefore the scope of the invention is not to beconstrued as limited to the preferred embodiment.

The invention claimed is:
 1. A process of preparing N-[4-[[4-(diethylamino)phenyl](2,5-disulfophenyl)methylene]-2,5-cyclohexadien-1-ylidene]-N-ethylethanaminium,sodium salt comprising combining a suspension of isoleuco acid of theformula

in a polar solvent with silver oxide, recovering isosulfan blue acid,and treating the isosulfan blue acid with a sodium solution.
 2. Theprocess according to claim 1 comprising sulfonation of2-chlorobenzaldehyde to obtain 2-chlorobenzaldehyde-5-sulfonic acidsodium salt of the formula

followed by nucleophilic displacement of the chloride in2-chlorobenzaldehyde-5-sulfonic acid sodium salt with an alkali metalsulfite and bisulfite to obtain benzaldehyde-2,5-disulfonic acid,disodium salt of the formula

and condensing the benzaldehyde-2,5-disulfonic acid, disodium salt ofthe formula (3) with N, N-diethylaniline using urea and glacial aceticacid to provide isoleuco acid of the formula (4).
 3. The process ofpreparing 2-chlorobenzaldehyde-5-sulfonic acid, sodium salt of formula(2) according to claim 2 comprising reacting 2-chlorobenzaldehyde withsulfuric acid.
 4. The process according to claim 1 wherein the polarsolvent is methanol.
 5. The process according to claim 2 of preparingfree benzaldehyde-2,5-disulfonic acid, di-sodium salt of the formula

wherein the alkali metal sulfite and bisulfite comprise sodium sulfiteand sodium bisulfite salts.
 6. The process according to claim 5 whereinthe reaction is carried out in a pressure vessel at 170-180° C. for 5 to7 hours.
 7. The process according to claim 6 wherein the reaction iscarried out under a pressure of 140 to 150 psi.
 8. The process accordingto claim 2 comprising precipitating inorganic salts which will hinderthe rate of reaction using methanol or one or more C₁₋₄ lower alcohols.9. The process according to claim 2 in which thebenzaldehyde-2,5-disulfonic acid disodium salt is purified by extractingwith a non-aqueous polar solvent followed by its precipitation in ahalogenated or non-halogenated non-polar solvent which is miscible withthe non-aqueous polar solvent.
 10. The process according to claim 9wherein the nonaqueous polar solvent is N,N dimethylformamide and thenonpolar solvent is dichloromethane.
 11. The process according to claim1 wherein the isoleuco acid of the formula (4) is prepared by combininga benzaldehyde-2,5-disulfonic acid, disodium salt of the formula

with N,N-diethylaniline, and urea and glacial acetic acid.
 12. Theprocess according to claim 11 performed at reflux conditions for 20-25hours at 115 to 120° C.
 13. The process according to claim 11 comprisingprecipitating a crude solid using methanol or a C₁₋₄ lower alcohol. 14.The process according to claim 11 in which the crude solid is furtherpurified using water.
 15. The process according to claim 1 comprisingoxidation of isoleuco acid of the formula (4) with silver oxide inmethanol to obtain a reaction mass.
 16. The process according to claim15 comprising stirring the reaction mass for 12-14 hours, and filteringthe silver oxide to provide a filtrate.
 17. The process according toclaim 16 comprising passing the filtrate through a bed of silica gel andcelite and passing the filtrate through a zeolite bed optionally treatedwith an acid or base.
 18. The process according to claim 17 furthercomprising passing the filtrate through a 0.2 micron filtration unit.19. The process according to claim 16 comprising precipitating thefiltrate using a non-polar solvent miscible with the filtrate.
 20. Theprocess according to claim 19 wherein the non-polar solvent is isopropylether.
 21. The process according to claim 1 comprising adjusting theN-[4-[[4-(diethylamino)phenyl](2,5-disulfophenyl)methylene]-2,5-cyclohexadien-1-ylidene]-N-ethylethanaminiumto a pH greater than 6.0 using an aqueous inorganic or organicderivative of sodium or a combination thereof.
 22. The process accordingto claim 21 wherein the pH is adjusted using sodium bicarbonatesolution.
 23. The process according to claim 1 comprisingrecrystallization ofN-[4-[[4-(diethylamino)phenyl](2,5-disulfophenyl)methylene]-2,5-cyclohexadien-1-ylidene]-N-ethylethanaminiumusing a solvent selected from the group consisting of a polar solvent, anon-polar solvent and a combination thereof to afford HPLC puritygreater than 99.5%.